Chromosome abnormalities are often associated with genetic disorders, degenerative diseases, and cancer. Chromosomal abnormalities can be of several types, including extra or missing individual chromosomes, extra or missing portions of a chromosome, breaks and chromosomal rearrangements Chromosomal rearrangements include translocations (transfer of a piece from one chromosome onto another chromosome), dicentrics (chromosomes with two centromeres), inversions (reversal in polarity of a chromosomal segment), insertions, amplifications, and deletions.
The detection of chromosomal abnormalities in cells is important for many reasons, not the least being the detection of chromosomal abnormalities for prenatal and pre-implantation genetic diagnosis, and the determination of the karyotype of some cancers.
Prenatal diagnosis involves the genetic testing of foetal material. Typically, this involves removal of amniotic fluid surrounding the foetus and the analysis of cells in the fluid for chromosomal abnormalities. Prenatal diagnosis is important for the detection of foetuses that have significant chromosomal errors. Detectable chromosomal abnormalities occur with a frequency of approximately one in every 250 human births, and abnormalities that involve deletions or additions of chromosomal material often lead to foetal death or to serious mental and physical defects. For example, Down Syndrome can be caused by having three copies of chromosome 21 instead of the normal two copies, or by a segmental duplication of a subregion on chromosome 21.
Pre-implantation genetic diagnosis (PGD) involves the testing of genetic material from an embryo or an egg (oocyte) prior to implantation. Typically, this process involves the removal and analysis of one or more cells from an embryo fertilized in vitro, in order to determine if the embryo is suitable for implantation. In the case of maternally derived chromosomal abnormalities, the polar body from an oocyte can also be removed and the presence of a chromosomal abnormality detected.
Pre-implantation genetic diagnosis for chromosomal abnormalities is important for detecting embryos or oocytes that are suitable for implantation. Early human embryos have a very high frequency of chromosomal errors including aneuploidy, polyploidy and mosaicism, and it is likely that these chromosomal errors are responsible for the significant rate of implantation failure of in vitro fertilized embryos. In addition, where there is a possibility that an embryo or oocyte may contain a known chromosomal abnormality inherited from one of the parents, pre-implantation diagnosis can also be performed to select embryos or oocytes that do not have the known chromosomal abnormality.
The deletion or multiplication of copies of whole chromosomes or chromosomal segments also often occurs in cancerous cells and in many cases these chromosomal abnormalities contribute to the cells acquiring a cancerous phenotype. The detection of such chromosomal abnormalities is not only important for understanding the genetic basis of how some cells progress from a non-cancerous state to a cancerous state, but in some cases may provide useful information as to the diagnosis and treatment of a specific cancer.
Traditionally, cytogenetic or fluorescence in situ hybridization (FISH) techniques have been used for detecting chromosomal abnormalities. However, comparative genomic hybridization (CGH) now provides a powerful method to overcome many of the limitations of the traditional cytogenetic and FISH approaches. CGH involves the comparative, multi-colour hybridization of a reference nucleic acid population labelled in one fluorescent colour and a sample nucleic acid population labelled in a second fluorescent colour to all or part of a reference genome, such as a human metaphase chromosome spread. Comparison of the resulting fluorescence intensity at locations in the reference genome permits determination of the copy number of chromosomal sequences in the sample population.
Although CGH has provided an improvement over traditional cytogenetic and FISH technologies, there are still many deficiencies associated with CGH, including the length of time required to perform the analysis. For example, standard CGH with metaphase spreads takes at least 72 hours to complete, and if PGD is being performed on a single cell taken from an embryo, then embryo cryopreservation before implantation must be performed so as to allow sufficient time to complete the procedure.
The present invention relates to the identification of an improved method for performing comparative genomic hybridization and to nucleic acid arrays suitable for comparative genomic hybridization.
Throughout this specification reference may be made to documents for the purpose of describing various aspects of the invention. However, no admission is made that any reference cited in this specification constitutes prior art in particular, it will be understood that the reference to any document herein does not constitute an admission that any of these documents forms part of the common general knowledge in the art in Australia or in any other country. The discussion of the references states what their authors assert, and the applicant reserves the right to challenge the accuracy and pertinency of any of the documents cited herein.